Esquire 3000 plus mass spectrometer

Mass spectrometry data were obtained in the positive ion mode using an Esquire 3000 Plus mass spectrometer (Bruker Daltonics), coupled with the HPLC instrument. The eluent was the same used in the HPLC analysis with the addition of 2% (v/v) formic acid to solvent A. Samples were introduced into the electrospray source at 70 μL/min. Capillary voltage was set at 4 kV, with a temperature of 250 °C. The nebulizer, dry gas, and dry temperature were set at 20 psi, 7.0 L/min, and 325 °C, respectively.

The phenolic compounds of A. nordenskioeldii were extracted and analysed according to [11 (link)]. Briefly, 20% ethanol extracts were characterized by liquid chromatography mass spectrometry (LC-MS) with an Agilent ChemStation 1100, equipped with a Synergy Hydro column 4 µm column (Phenomenex) and a diode array detector set at 280 nm, coupled with an Esquire 3000plus mass spectrometer with electrospray ionization (Bruker Daltonics, Bremen, Germany).

Phenolic compound analyses were carried out using a 1290 Infinity UPLC (Agilent Technologies, Les Ulis, France). The UPLC system was coupled to an Esquire 3000 plus mass spectrometer from Bruker Daltonics (Wissembourg, France). Five µL was injected into a column Zorbax SB-C18 (2.1 × 100 mm, 1.8 µm) (Agilent Technologies, Les Ulis, France). Two different solvents were used as a mobile phase: solvent A (water/formic acid 99.9:0.1, v/v) and solvent B (acetonitrile/formic acid 99.9:0.1, v/v), at a flow rate of 0.4 mL/min and a gradient as follows in solvent A: 0 min 1% B, 0.4 min 1% B, 2 min 10% B, 6 min 35% B, 7 min 50% B, 8.8 min 70% B, 10.8 min 92% B, 11 min 100% B, 12 min 100% B, 12.2 min 1% B, and 15.2 min 1% B. The MS/MS parameters were set as follows: negative mode; capillary tension +4000 V; nebulizer 40 psi; dry gas 10 L/min; dry temperature 365 °C; and scan range m/z 100 to 1400. Data were processed using HyStar 3.2 software (Bruker Daltonics, Wissembourg, France).

Phenolic compounds analyses were carried out using a 1290 Infinity UPLC from Agilent Technologies. The UPLC system was coupled to an Esquire 3000 plus mass spectrometer from Bruker Daltonics (Wissembourg, France). 5 μL were injected into a Zorbax SB-C18 column (2.1 × 100 mm, 1.8 μm) from Agilent Technologies. Two different solvents were used as a mobile phase: solvent A (water/formic acid 99.9:0.1 v/v) and solvent B (acetonitrile/formic acid 99.9:0.1 v/v), at a flow rate of 0.4 mL/min and a gradient as follows in solvent A: 0 min 1% B, 0.4 min 1% B, 2 min 10% B, 6 min 35% B.7 min 50% B, 8.8 min 70% B, 10.8 min 92% B, 11 min 100% B, 12 min 100% B, 12.2 min 1% B, 15.2 min 1% B. The MS/MS parameters were set as follows: negative mode; capillary tension +4000 V; nebulizer 40 psi; dry gas 10 L/min; dry temperature 365 °C; and scan range m/z 100 to 1400. Data were processed using HyStar 3.2 software (Bruker Daltonics).

¹H NMR, ¹³C NMR, and DEPT spectra were recorded at 500 MHz for ¹H and at 125 MHz for ¹³C, with a Bruker DRX 500 instrument in DMSO-d₆ solution. ESI-MS spectra were recorded on a Bruker Esquire 3000^plus mass spectrometer, using a negative ion probe with samples dissolved in methanol.
1,1-Diphenyl-2-picrylhydrazyl (DPPH), ferrozine, and Folin-Ciocalteu reagent were purchased from Sigma Aldrich Co (St. Louis, MO, USA). The reference standard Pycnogenol (Maritime Pine Extract, containing 673 mg proanthocyanidins per g material) was purchased from the United States Pharmacopeial Convention (Rockville, MD, USA). The reference standards gallic acid (purity ≥ 98%), rutin (purity ≥ 98%), proanthocyanidins (purity ≥ 95%), and ascorbic acid (purity ≥ 99%) were purchased from Internet Aladdin reagent database Inc. (Shanghai, China). The detection kits for protein content, total antioxidative capacity (T-AOC), superoxide anion free radical, hydroxyl free radical and malondialdehyde (MDA), aspartate aminotransferase (AST), alanine aminotransferase (ALT), superoxide dismutase (SOD), and reduced glutathione (GSH) were purchased from Nanjing Jiancheng Bioengineering Institute (Nanjing, Jiangsu, China). All other reagents were of analytical grade and made in China.

Glycopeptide mixtures were separated by the Prominence HPLC system (Shimazu, Kyoto, Japan) using a PEGASIL ODS 3 μm 1 mm i.d. × 100 mm column, and by the mobile phase of 0.1% formic acid (FA) and acetonitrile containing 0.1% FA with linear gradient mode at room temperature. The flow rate was 50 μL/min, and the detection was performed on an Esquire 3000 plus mass spectrometer equipped with an electrospray ionization interface (Bruker Daltonics GmbH, Bremen, Germany). The mass spectrometric parameters were as follows: polarity = positive ion mode, ion source gas (nitrogen) pressure = 10 psi, dry gas = 4.0 L/min, dry temperature = 250 °C and scan range = from 400 to 2000 m/z. In MS/MS experiments, the precursor isolation window was set to ±2 Da, and He was used as the collision gas. The operation was carried out on trapControl software and data were acquired and processed with Compass DataAnalysis (Bruker Daltonics GmbH, Bremen, Germany).